Proteolytic activity of cultured Pseudoperkinsus tapetis extracellular products

Several pathogenic protozoan release proteases are necessary for host invas ion and initiation of infection. We have identified proteolytic activities in extracellular proteins secreted by the clam parasite Pseudoperkinsus tap etis (Mesomycetozoa) in vitro. The protein concentration of the P. tapetis extracellular products (ECP) increased only during the first week of cultur e. The appearance of new proteins of 10 and 157 kDa at the second week samp le and of 12 kDa at the third week sample was shown by SDS-PAGE. The protea se activity rapidly increased in the first 3 weeks of culture, and five cle ar bands of 23, 29, 60, 67 and 96 kDa with proteolytic activity were detect ed in the ECP on gelatin SDS-PAGE. Using inhibitors, the proteases were ide ntified as members of the Ca2+ dependent, serine protease family. Their opt imum pH was higher than pH 9.4. The protease activity of the P. tapetis ECP was different than that described for Perkinsus marinus, an oyster pathoge n very similar morphologically to the clam parasite and member of the genus in which P. tapetis had been initially included. (C) 2001 Elsevier Science Inc. All rights reserved.